1. Field of the Invention
This invention relates to an imaging method for a scanning microscope wherein an image of a sample is formed with an optical type scanning microscope. This invention particularly relates to an imaging method for a scanning microscope wherein the image quality of an image of a sample is prevented from becoming bad due to a deviation in position of a light beam which has passed through the sample. This invention also relates to a confocal scanning microscope which is of a transmission type and in which the image quality of an image of a sample is not adversely affected when the position, at which an image of a light beam which has passed through the sample is formed, deviates in directions normal to the optical axis of an optical means.
2. Description of the Prior Art
Optical type scanning microscopes have heretofore been used. With the scanning microscope, a light beam is converged to a small light spot on a sample, and the sample is two-dimensionally scanned with the light spot. The light beam, which has passed through the sample during the scanning, or the light beam, which has been reflected from the sample during the scanning, is detected by a photodetector. An enlarged image of the sample is thereby obtained. An example of the scanning microscope is disclosed in Japanese Unexamined Patent Publication No. 62(1987)-217218.
In the conventional optical type scanning microscopes, a mechanism which two-dimensionally deflects a light beam by a light deflector are primarily employed. Also, in U.S. patent application Ser. No. 587,122, a novel mechanism has been proposed wherein a light projecting optical means and a light receiving optical means are supported on a single movable member, the movable member is moved with respect to a sample supporting member, and a light spot of a light beam is thereby caused to scan a sample.
With transmission type scanning microscopes wherein a light beam, which has passed through a sample, is detected, the problem occurs in that the contrast and resolution of a microscope image become bad due to a deviation in position of the light beam, which has passed through the sample. Specifically, in general, in the transmission type scanning microscopes, a pinhole plate is located on the upstream side of a photodetector in order to block a halo occurring with the light beam, which has passed through the sample, and light scattered from the sample. The light beam, which has passed through a pinhole of the pinhole plate is detected by the photodetector. However, if the light beam is refracted by the sample, the center point of the light beam, which has passed through the sample, deviates from the center point of the pinhole. Therefore, the light beam, which has passed through the sample, is eclipsed by the pinhole plate.
In order for the aforesaid problem to be eliminated, it is considered to move the pinhole plate in synchronization with the scanning of the light beam such that the deviation in position of the light beam, which has passed through the sample, may be compensated for. However, the extent to which the light beam is refracted by the sample varies for different samples. Also, even for a single sample, the extent to which the light beam is refracted by the sample varies for different parts of the sample in accordance with the optical properties of the parts. Therefore, actually, it is difficult to know in advance how the pinhole plate is to be moved.
Therefore, it may be considered that a deviation in position of the light beam, which has passed through the sample, be detected and, at the same time, movement of the pinhole plate be controlled in accordance with the results of the detection. However, the response speed of a control means is limited. Therefore, with such a method, the speed with which the light beam scans must be decreased in accordance with the response speed of the control means. Accordingly, a long time is required for the imaging to be carried out, and a microscope image cannot be outputted with a rate on the order of the ordinary television rate.